Allergen-mediated activation of IgE-bound mast cells via high-affinity receptors (Fc?) plays an important role in allergic diseases. In the traditiona view, binding of monomeric IgE to Fc? before exposure to antigen is a passive sensitization step, which does not cause mast cell activation. However, two studies including ours in 2001 demonstrated that binding of monomeric IgE to Fc? in the absence of antigen promotes mast cell survival. Subsequently, we found an extensive heterogeneity among IgE molecules in their ability to influence the activity of mast cells via Fc?: in a broad spectrum of heterogeneity, so called highly cytokinergic (HC) IgEs can induce all kinds of activation events in the absence of specific antigen, including strong survival promotion, degranulation, production of leukotrienes and cytokines, chemotaxis, etc., whereas poorly cytokinergic (PC) IgEs can induce only weak survival promotion. More recently, we showed that a subset of IgE molecules bind directly to histamine-releasing factor (HRF), a molecule secreted during the late phase of allergic reactions that can promote allergic reactions in several allergy models. Anaphylaxis is a potentially life- threatening, severe allergic reaction that is rapid in onset and is often caused by foods. In a well-established mouse model, food allergy-induced diarrhea depends on IgE, Fc?, and mast cells. In our preliminary experiments, we observed contrasting outcomes of allergen intake, i.e., severe (early onset and even deaths) vs. mild (late onset and less severe diarrhea) anaphylactic reactions, in genetically identical inbred BALB/c mice using the same allergen and the same procedures. Under these conditions, IgE molecules would be generated in individual mice in a rather stochastic manner in terms of the usage and combinatorial joining of V, D, and J gene segments and the pairing of heavy and light chains. Therefore, we hypothesize that the differing outcomes of allergic reactions derive from the heterogeneity of IgE molecules generated in individual mice. Mice that exhibit severe anaphylactic responses in food allergy could express HC IgEs with strong Fc?- cross-linking capacity and/or HRF-reactive IgEs, which should be able to cross-link Fc? together with HRF. To test this hypothesis, (i) we will analyze the paired IgE heavy and light chain sequences in IgE+ B cells to find distinct structural features in IgE molecules between severely and mildly anaphylactic mice. Using recombinant IgE molecules derived from individual IgE+ B cells, (ii) we will investigate their abilities to exhibit HC vs. PC property, to bind HRF, and to induce food allergy. Through this project, we will likely show that IgE heterogeneity plays a critical role in determining the severity or onset of food allergy-induced anaphylaxis. Such results will prompt clinical studies to search for a biomarker(s) to detect IgE heterogeneity as well as to identify patients who may or may not respond well to allergen-specific immunotherapy and/or anti-IgE therapy.